Purpose: [13C1] Pyruvate metabolites in a hyperpolarized state have been visualized in whole rodent liver in vivo and ex-vivo. However, this approach does not permit the necessary radiology-histopathology needed to adequately evaluate the technique as a marker of the relevant biologic process. Precision cut liver slices have the potential to breach this obstacle. We aimed to validate the use of precision cut liver slices for hyperpolarized metabolic investigation in a mouse model. Following validation of the slices viability by 31P NMR spectroscopy and detection of ATP signals, hyperpolarized [13C1] pyruvate was administered to the slices and its metabolism was followed.
Materials and Methods: Fresh normal livers were harvested from five mice. The livers were sliced with a microtome-tissue slicer to 500 μm thickness slices and placed in an ice-cold recovery medium. The slices were then placed in an NMR spectrometer (5.8 T, RS2D, France) and perfused continuously with oxygenated growth media at 37 °C. 31P spectra were acquired to evaluate the presence of ATP. After ATP production was observed, hyperpolarized [13C1]pyruvate was flushed into the NMR tube in the spectrometer, into the liver. Consecutive 13C NMR spectra were acquired immediately after the injection. Spectral analysis was performed using MNova (Mestrelab Research, Santiago de Compostela, Spain).
Results: The 31P spectra collected from each liver showed the characteristic signals of ATP, confirming the viability of the tissues slices in the spectrometer. After each of the [13C1] pyruvate injections, both [13C1] lactate and [13C1] alanine signals were detected within 20 sec, for a duration of 1.5 min.
Conclusion: We were able to maintain and show the viability of precision-cut mouse liver slices in an NMR spectrometer as well as record hyperpolarized [13C1] pyruvate metabolism for the first time. This suggests that it can be used for ex-vivo evaluation of liver metabolism. This strategy is translational to the study of human liver metabolism ex-vivo as very small amounts of tissue are required (less than 1 gram). This approach is likely to lead to more accurate assessment of intracellular processes in a personalized manner.
